To date, biosensors have generally used optical measurement methods utilizing optical dyes. Biosensors using optical measurement methods have advantages of considerably excellent sensitivity and superior sensing selectivity, but disadvantages of high-cost measurement apparatuses and long measurement time due to the necessity of pre-treating processes such as adhesion of optical dyes to reagents. In an attempt to solve these disadvantages, a great deal of research is being actively conducted on nano-biosensors. Nano-biosensors are manufactured based on nanotechnology. Nanomaterials such as nanowires or nanoparticles have a considerably large surface area per unit volume, linkable to bio-molecules, thus exhibiting considerably superior sensitivity, as compared to conventional micromaterials. Furthermore, biosensors using materials such as optical dyes indirectly measure biomaterials through light for a long time, whereas nanobiosensors can advantageously rapidly real-time monitor sensitivity through direct binding of nanomaterials to target biomaterials.
Research associated with nanomaterials for biosensors is actively conducted based on carbon nanotubes, and metal and inorganic semiconductor nanomaterials. Carbon nanotubes are continuously researched by development of various synthesis methods, but have not been put into practical application due to disadvantages such as high manufacturing costs, chirality-dependent electrical properties and inactive surfaces. In addition, metal and inorganic semiconductor nanomaterials are limited in terms of biocompatibility due to toxicity. On the other hand, conductive polymers have various advantages such as various molecule designs, easy processing, low-weight and flexibility (Polymer Science and Technology, vol. 18, pp. 306-310, 2007).
The oxidation level of conductive polymers can be readily controlled by chemical or electrochemical doping/dedoping. This induces sensitive rapid reactions (variations in electrical conductivity or color) with specific chemical and biological species (See: Chem. Rev., vol. 100, pp. 2537-2574, 2000). This property enables application of conductive polymers to various sensor activities. In practical use, conductive polymers are known to be more sensitive to external environmental variations than other sensing materials due to inherent transfer properties such as such as electrical conductivity and energy transfer (See: Acc. Chem. Res., vol. 31, pp. 201-207, 1998). In addition, one-dimensional conductive polymer nanostructures such as nanorods, nanofibers and nanotubes have a larger surface area, thus providing sensitivity and real-time reactions amplified by increased interaction with analytes (See: Nano Lett., vol. 4, pp. 491-496,2004; Nano Lett., vol 4, pp. 671-675, 2004). In spite of these advantages, the absence of reproducible and reliable methods for preparing nanoparticles has hampered development of sensors using conductive polymer nanostructures. In particular, methods for fabricating conductive polymer nanomaterials have been limited to methods using expensive templates such as porous alumina membranes or polycarbonate membranes, and have a serious disadvantage of considerably low yield through complicated multi-step synthesis (See: Chem. Mater., vol. 8, pp. 2382-2390; Science, vol. 296, pp. 1997).
In general biosensors, for reactions selective to specific target materials, receptors, RNA/DNA aptamers, proteins, or the like are incorporated into transducers. Receptors are generally adhered to the surface of transducers through absorption, entrapment and covalent bonding. Of these, the method for adhering receptors through covalent bonding has an advantage of considerably superior chemical and physical stability as compared to other methods. However, this method requires chemical functional groups applicable to the surface of transducers. Metal, inorganic semiconductor and polymer nanomaterials as well as carbon nanotubes have inactive surfaces, thus requiring additional surface-treatment processes to incorporate surface functional groups.
Probing and detection of biomaterials are currently carried out in solution. For this purpose, carbon nanotubes and inorganic semiconductor nanomaterials are directly fixed on an electrode surface by photolithography or electron-beam lithography. However, conductive polymer nanomaterials are disadvantageously unsuitable for lithography due to the risk of chemical and physical damage. In addition, most conductive polymers exhibit low adhesion force to an electrode substrate made of a material such as silicon, glass, or metal. Due to these disadvantages, development of biosensors using conductive polymer nanostructures has been considerably limited.
The recently discovered vascular endothelial growth factor (hereinafter, referred to as “VEGF”) is an endothelial cell-specific mitosis accelerant known as an important inducing factor which mediates generation mechanisms and formation of blood vessels under physiological or pathological conditions and creates new blood vessels in a variety of tumors. It has already been demonstrated that such blood vessel formation is related to pathogenesis of various diseases. Such diseases include ocular neovascular syndrome, rheumatoid arthritis and psoriasis such as solid tumors, proliferative retinopathy or age-related macular degeneration (AMD) (Documents ([Folkman et al. J. Biol. Chem. 267:10931-10934 (1992)]; [Klagsbrun et al. Annu Rev. Physiol. 53:217-239 (1991)]; and [Garner A, Vascular diseases. In: Pathobiology of ocular disease: a dynamic approach. Garner A, Klintworth G K, Eds. 2nd Edition (Marcel Dekker, NY, 1994), pp 1625-1710])). For solid tumors, creation of new blood vessels causes tumor cells to exhibit superior development superiority and proliferation autonomy, as compared to normal cells. Accordingly, for breast cancers and other tumors, correlation between microvessel density of the tumor sites and patient survival rate was reported (Documents ([Weidner et al. N Engl J Med 324:1-6 (1991)]; [Horak et al. Lancet 340:1120-1124 (1992)]; and [Macchiarini et al. Lancet 340:145-146 (1992)])). As such, VEGF plays an essential role in development of various tumor cells and is detected in large amounts where tumors are created. Accordingly, accurate sites and rapid diagnosis of formed tumors can be realized by detecting VEGF amount variations. Biosensors for detecting various VEGFs have been developed to date (Biosens Bioelectron 2009; 24:1801-05). However, to date, there is a need for continuous development of biosensors due to disadvantages such as limited detection concentration and long detection time.
Accordingly, for industrial application of basic technologies, there is an increasing need for technologies for efficiently and simply controlling diameters of conductive polymer nanomaterials having surface chemical functional groups and technologies for fabricating high-performance biosensors based on the same.